Spermatogenesis is a complex developmental process where undifferentiated spermatogonia are differentiated into mature spermatozoa that are then able to fertilize oocytes. In mammals, this process involves hundreds of genes that are differentially regulated in spermatogenic tissue at particular stages of sperm differentiation. Identifying critical, regulatory genes controlling spermatogenesis can be significantly Informed through comparative gene expression and functional studies utilizing simpler animal models that have conserved developmental processes. My proposed research will characterize gene expression, protein-protein interactions, and gene silencing coupled with transcriptional profiling to elucidate the integral role of three orphan nuclear receptors (testicular orphan receptors and germ cell nuclear factor) in sperm development and differentiation utilizing the emerging model Nematostella vectensis (starlet sea anemone). Each of these receptors has been shown to be highly expressed in mammal testes at discrete stages of sperm differentiation;thus results from the proposed research will characterize novel roles for nuclear receptor function related to human male fertility. The proposed research supports the mission of the Reproductive Sciences Branch of The Eunice Kennedy Shriver National Institute of Child Health and Human Development by identifying genetic mechanisms that underlie reproductive health in humans. In addition, these three transcription factors, like other genes in the nuclear receptor superfamily, have pleiotropic roles in embryogenesis including roles in development of the nervous and cardiovascular system and in cancer due to expression in various embryonal carcinoma cell lines. I will take advantage of the experimental tractability and tools already developed for Nematostella to explore the additional roles of these receptors in embryo development and abnormal cell regulation. Relevance to Public Health Identifying and characterizing genes that play a fundamental role in human fertility is a critical step towards advancing diagnosis and treatment in human reproduction. Utilizing morphologically simple and experimentally tractable animal models to test specific molecular processes can result in rapid progress in understanding mechanisms. Nematostella, an emerging model system, will be used to study the role of three genes highly expressed and functionally important in male spermatogenesis to understand their role in differentiation of mature sperm from undifferentiated stem cells.